Insulin has profound effects on a number of diverse cellular processes: protein synthesis (1), glycogen and lipid metabolism (2,3), glycose, amino acid and ion transport (4,5) and cell growth (6). It produces these immediate and delayed effects subsequent to its interaction with a specific receptor on the cell membrane. Insulin action involves: (1) interaction of the hormone with a receptor site upon the cell surface; (b) signal(s) generated by the hormone-receptor interaction at the cell surface and inside the cell; (c) physiological changes in cell function. The coupling mechanism(s) between the insulin-receptor interaction and the subsequent metabolic effects produced by insulin action remains unclear. We are studying the coupling mechanisms by exploiting our recent findings that (1) insulin action on fat cells increases the phosphorylation of ATP-citrate lyase; (2) this increase in enzyme phosphorylation is not mediated by cyclic AMP-dependent protein kinase in vivo; (3) ATP-citrate lyase is phosphorylated at a minimum of two sites, at least one of which is structural and another catalytic; and (4) structural site phosphorylation is hormone dependent. The hypothesis is: Does insulin action increase protein kinase activity which increases the phosphorylation (from ATP) of structural phosphates of ATP-citrate lyase? The Specific Aims are (1) Isolation of a non-cyclic AMP dependent protein kinase which phosphorylates the structural phosphates of ATP-citrate lyase; (2) study of the control mechanisms for this protein kinase; (3) study of the relationship of enzyme phosphorylation to enzyme activity during different physiological states; (4) study of the catalytic and structural phosphorylation site of ATP-citrate lyase. Employing these same methodologies we intend to purify and identify the complex of proteins, the phosphorylations of which are markedly affected by insulin and beta-agonist actions in a reciprocal manner (bands 5, 6a, 6b, Subunit Mr 65,000-70,000, 65,000 and 62,000 respectively).